The process by which Treg cells develop within the thymus is poorly understood. During the last grant period, our laboratory established that thymic expression of a functional IL-2R was one critical step for the thymic production of Treg cells. A key role of IL-2for Treg cells is not just limited to the thymus as we further showed that the IL-2/IL-2R interaction was essential for extensive expansion of Treg cells in neonatal lymph nodes, leading to a normal complement of these cells in secondary lymphoid tissue. Given the important role for Treg cells in T cell tolerance, understanding the basis by which this T cell subset develops may shed light on the basis of autoimmune disease and facilitate application of these cells in clinical situations, from autoimmunity, to organ transplantation, and tumor immunity. In this regard, a number of important unanswered questions remain concerning the contribution of IL-2to the production of Treg cells. These include: What are the consequences of IL-2 signaling for Treg cells within the thymus? Is IL-2just a growth/survival factor or does it more broadly program Treg cells? Is the role of IL-2similar in both the thymus and in neonatal secondary lymphoid tissue? What are the signaling requirements through the IL-2R for the production of Treg cells? What is the cellular source for IL-2for Treg cells? The major objective of this competitive renewal is to answer these questions and hence establish a much more detailed understanding concerning the contribution of IL-2in the developmental production of Treg cells. The specific aims are:1) To define the consequences of the IL-2/IL-2R interaction in Treg cells during thymic development; 2) to investigate the requirements for IL-2R signaling for the production of Treg cells in vivo: and 3) to determine the basis for IL-2-dependent Treg proliferation within neonatal lymph nodes and directly identify the keyIL-2 producing cell.